Virulence and Molecular Diversity in Cochliobolus sativus

ABSTRACT Spot blotch, caused by the fungal pathogen Cochliobolus sativus, is an important disease of barley in many production areas of the world. To assess genetic diversity in this pathogen, a worldwide collection of C. sativus isolates was evaluated for virulence on barley and DNA polymorphism. Three pathotypes (0, 1, and 2) were identified among the 22 isolates tested in this study and the 36 isolates characterized previously on three barley differentials (ND5883, Bowman, and NDB112) that differ in their resistance to C. sativus. Pathotype 2, which exhibits high virulence on cv. Bowman, was only found in North Dakota, whereas the other two pathotypes occurred in many other regions of the world. Genetic diversity of the 58 C. sativus isolates, together with isolates of three related pathogenic Cochliobolus spp. (C. heterostrophus, C. carbonum, and C. victoriae) was analyzed using amplified fragment length polymorphism (AFLP) markers. A total of 577 polymorphic AFLP markers were recorded among the 70 isolates of the four Cochliobolus spp. using eight primer combinations. Cluster analysis revealed distinct groups corresponding to the four different species, except in one case where race 0 of C. carbonum was placed in an outgroup that may belong to a different species. In C. sativus, 95 polymorphic AFLP markers were detected with the eight primer pairs used, and each isolate exhibited a unique AFLP pattern. Allelic diversity in the pathotype 2 group was lower (0.10) than in the pathotype 0 (0.23) and pathotype 1 (0.15) groups, indicating that pathotype 2 may have arisen more recently. Cluster analysis did not reveal a close correlation between pathotypes and AFLP groups, although two AFLP markers unique to pathotype 2 isolates were identified. This low correlation suggests that genetic exchange may have occurred through parasexual recombination in the fungal population. Some isolates collected from different regions of the world were clustered into the same AFLP group, suggesting that migration of the fungal pathogen around these regions has occurred.     

Isolation and Characterization of an Endopolygalacturonase from Cochliobolus sativus and a Cytological Study of Fungal Penetration of Barley

ABSTRACT Endopolygalacturonase (EPG) of Cochliobolus sativus was produced in shake culture and purified by high-performance liquid chromatography. The enzyme had a molecular mass of 34,000 Da, an isoelectric point in the range of 9.0 to 9.5, exhibited endo activity, was nongly-cosylated, and was inhibited by polygalacturonase-inhibiting proteins from bean, pear, and tomato. The amino terminus contained a 14 amino acid region homologous to a region at the N terminus of an EPG of C. carbonum. C. sativus EPG-specific monoclonal antibodies (MAbs) were generated. Western blot analysis confirmed the specificity of the antibodies for the EPG and detected the enzyme in an extract from Hordeum vulgare (cv. Golden Promise) leaf segments infected with C. sativus. Using conventional immunogold and enzyme-gold cytochemical methods, homogalacturonan, esterified pectin, and cellulose were localized in healthy and infected barley leaf epidermis at the electron microscope level. Additionally, the leaf cell wall polysaccharides recognized by purified C. sativus EPG were localized at the electron microscope level, using the purified enzyme as a primary cytochemical reagent, followed by a gold-labeled MAb specific for the enzyme. Loss of polygalacturonic acid in the vicinity of the invading pathogen was visualized cytochemically at the electron microscope level. These observations suggest the involvement of EPG during host penetration by the fungus.     

Structure of the Cochliobolus sativus population variability

The genetic diversity of several local populations of the fungal pathogen Cochliobolus sativus, collected over 3 years from different regions of the Czech Republic, was examined using amplified fragment length polymorphism (AFLP). A high level of variability was found even among isolates from one lesion. Measures of multilocus linkage disequilibrium suggested that recombination has a minor impact on the genetic structure of populations. Cochliobolus sativus forms genetically divergent populations (F_ST = 0·33), indicating a low level of geneflow between populations. This was supported by a significant correlation between genetic diversity and geographical distances up to 80–100 km. The most likely explanation for the genetic variability is that the fungus forms conidia with highly variable chromosomal rearrangements. The differentiation observed among local populations implies that genetic drift, including a founder effect, combined with restricted migration generates the structure of C. sativus populations.     

A Compromised Mlo Pathway Affects the Response of Barley to the Necrotrophic Fungus Bipolaris sorokiniana (Teleomorph: Cochliobolus sativus) and Its Toxins

ABSTRACT In search of new durable disease resistance traits in barley to control leaf spot blotch disease caused by the necrotrophic fungus Bipolaris sorokiniana (teleomorph: Cochliobolus sativus), we developed macroscopic and microscopic scales to judge spot blotch disease development on barley. Infection of barley was associated with cell wall penetration and accumulation of hydrogen peroxide. The latter appeared to take place in cell wall swellings under fungal penetration attempts as well as during cell death provoked by the necrotrophic pathogen. Additionally, we tested the influence of a compromised Mlo pathway that confers broad resistance against powdery mildew fungus (Blumeria graminis f. sp. hordei). Powdery mildew-resistant genotypes with mutations at the Mlo locus (mlo genotypes) showed a higher sensitivity to infiltration of toxic culture filtrate of Bipolaris sorokiniana as compared with wild-type barley. Mutants defective in Ror, a gene required for mlo-specified powdery mildew resistance, were also more sensitive to Bipolaris sorokiniana toxins than wild-type barley but showed less symptoms than mlo5 parents. Fungal culture filtrates induced an H2O2 burst in all mutants, whereas wild-type (Mlo) barley was less sensitive. The results support the hypothesis that the barley Mlo gene product functions as a suppresser of cell death. Therefore, a compromised Mlo pathway is effective for control of biotrophic powdery mildew fungus but not for necrotrophic Bipolaris sorokiniana. We discuss the problem of finding resistance traits that are effective against both biotrophic and necrotrophic pathogens with emphasis on the role of the anti-oxidative system of plant cells.     

Polyoxin Resistance of Reddish Brown Laboratory Mutants of Cochliobolus heterostrophus

Six reddish brown polyxin-resistant mutants of Cochliobolus heterostrophus were isolated after ethyl methanesul-phonate and N-nitroquinoline oxide mutageneses followed by selection on polyoxin. All the mutants were highly resistant to polyoxin (MIC > 1600 μg/ml). When mutants were crossed with the wild-type strain, all crosses had a 1 : 1 ratio of mutant (reddish brown pigmentation and polyoxin resistance) : wild type (non-reddish brown pigmentation and polyoxin sensitivity), indicating that the phenotypes in these strains were due to alteration at a single gene locus in each strain. Allelism tests revealed the existence of two loci, Pol2 and Pol5. The results of the crossing and mutation-rate studies suggest that the each gene was pleiotropic for the reddish brown color and polyoxin resistance. Received 19 September 2001/ Accepted in revised form 25 December 2001     

Durability of Resistance and Cost of Virulence

A seasonal model, where a growing season is defined as the time between sowing and harvest and alternates with an inter-crop period, was derived to study the effects of the ‘cost of virulence’ and cropping ratio on durability of resistance. We assumed a single strain of virulent pathogen, a single strain of avirulent pathogen and two cultivars (one resistant and one susceptible) and studied two measures of durability of resistance (‘take-over time’ and ‘usefulness time’). Take-over time is defined as the time needed for the virulent strain of the pathogen to reach a preset threshold and predominate over the previous pathogen population. Usefulness time is the time needed before the estimated gain in green canopy area duration per plant through the use of the resistant cultivar becomes negligible. The model suggested that, although it could take several seasons before the virulent strain of the pathogen predominated over the previous pathogen population, the usefulness time of the resistant cultivar was always much shorter. Furthermore, increasing selection for the virulent strain of the pathogen (through increasing the cropping ratio of the resistant cultivar) caused the virulent strain of the pathogen to invade the system more rapidly. Cost of virulence, reflecting differences in pathogen infection rates between the four possible combinations of cultivar/pathogen strain, significantly affected durability of resistance, with the dynamics of the virulent and avirulent strains ranging from a case where the virulent strain of the pathogen died out to a case where the virulent strain of the pathogen invaded the resident pathogen population. An intermediate state, where the system reached equilibrium and the virulent strain of the pathogen neither became predominant nor died out, was defined as ‘coexistence’ of both strains of the pathogen. Occurrence of coexistence was directly related to the cost of virulence since it did not occur when virulence of the pathogen did not have a fitness cost. Two methods to include cost of virulence in the model gave similar results in relation to the two measures of durability of resistance studied.     

Effect of growth stage on host sensitivity to helminthosporol toxin and susceptibility to Cochliobolus sativus causing spot blotch on wheat

Five wheat genotypes with differing field resistance to Cochliobolus sativus were tested in greenhouse conditions for their reaction to 1.0 mM helminthosporol (HL) toxin and for their resistance to the pathogen at the 4-leaf seedling, booting, anthesis completed and late milk grain stages. HL caused distinctly necrotic and chlorotic lesions only on the adult plants of highly susceptible genotypes Ciano T-79 and Sonalika. The level of toxin sensitivity and disease severity increased with adult plant growth stage, suggesting that susceptibility is related to plant physiology. However, genotype BL 1473 was found to be susceptible to the fungus but insensitive to the toxin, indicating the limitations of using the toxin as a screening tool for resistance. The results confirm earlier studies which showed a lack of correlation between resistance at the seedling stage and field observations. This underlines the importance of field screening for improving resistance to spot blotch in wheat.     

Mechanisms of Induced Resistance in Barley Against Drechslera teres

ABSTRACT Quantitative and qualitative histopathological methods and molecular analyses were used to study the mechanisms by which preinoculation with either of the nonbarley pathogens, Bipolaris maydis and Septoria nodorum, inhibited growth of Drechslera teres. Collectively, our data suggest that induced resistance is the principal mechanism responsible for impeding the pathogen. The enhancement of resistance in the host was primarily manifested during penetration by D. teres, and after penetration, where growth of D. teres ceased soon after development of infection vesicles. Thus, 24 h after pretreatment with B. maydis or S. nodorum, the penetration frequency from D. teres appressoria was reduced from 42.7% in the controls to 9.5 and 14.8%, respectively. The reductions were associated with increased formation of fluorescent papillae in induced cells (early defense reaction). The postpenetrational inhibition of D. teres completely stopped fungal growth and was apparently linked to an enhancement of multicellular hypersensitive responses in inducer-treated leaves (late defense reaction). Papillae formation and multicellular hypersensitive reactions were also observed in fully susceptible, noninduced control leaves, but they were inadequate to stop fungal progress. Northern blots from leaves treated with either inducer alone support the conclusion that induced resistance is involved in suppression of D. teres by increased formation of papillae and hypersensitive reactions. Thus, the blots showed strong expression of several defense response genes that are involved in these reactions in barley attacked by Erysiphe graminis f. sp. hordei.     

The buffering capacity of the natural mycoflora of rye leaves to infection by Cochliobolus sativus,and its susceptibility to benomyl

Different densities of saprophytic colonization of rye leaves were obtained in field experiments by spraying with benomyl or water. In 1972 and in 1973, inoculation withCochliobolus sativus, just after flowering, resulted in 60% less necrosis on water-sprayed leaves than on benomyl-sprayed leaves. At that time, the natural mycoflora of the water-sprayed leaves amounted to 10 000 and 3000 propagules per cm² leaf surface in 1972 and 1973, respectively. The benomyl treatment reduced the colonization to 1200 and to 400 propagules per cm² in 1972 and 1973, respectively, which implied an apparent reduction of the antagonistic capacity of the mycoflora. In 1974, the saprophytic colonization of the water-sprayed leaves reached only 500 propagules per cm² just after flowering, a population density not high enough to be antagonistic. Benomyl had a differential effect on the phyllosphere fungi:Cryptococcus spp. were not affected,Sporobolomyces roseus andCladosporium spp. were reduced to less than 6% of the control populations, andAureobasidium pullulans was eliminated. When later in the seasons of 1972 and 1974 the white yeasts on the benomyl-sprayed leaves reached population densities of 6500 propagules per cm² and more, inoculation of these leaves resulted in a necrosis similar to that of the water-sprayed leaves with higher population densities. Above a population density of 6500 propagules per cm² no correlation existed between the density of the antagonists and their action onC. sativus.Samenvatting In veldproeven werden roggebladeren, voor de bloei en op verschillende tijdstippen na de bloei, geïnoculeerd metCochliobolus sativus (Helminthosporium sativum) teneinde het antagonisme tegen dit pathogeen door de van nature voorkomende schimmelflora —inclusief gisten—te bestuderen. Eerder uitgevoerde kasproeven hebben aangetoond dat saprofytische fyllosfeerschimmels met succes met perthotrofe pathogene schimmels wedijveren om de voedingsstoffen die vrijkomen uit het stuifmeel (Warren, 1972b; Fokkema, 1973).Verschillende niveaus van saprofytische bladkolonisatie werden verkregen door de proefveldjes met benomyl of met water te behandelen (Fig. 1).C. sativus is relatief weinig gevoelig voor benomyl. Het gemiddeld percentage necrotisch bladoppervlak ten gevolge van inoculatie metC. sativus vlak na de bloei was in 1972 en in 1973 ruim 60% lager op met water behandelde bladeren dan op met benomyl behandelde bladeren (Fig. 2). Op dit moment heeft de natuurlijke schimmelflora op de met water bespoten bladeren een populatiedichtheid bereikt van 10000 propagula per cm² bladoppervlak in 1972 en van 3000 propagula per cm² in 1973. Benomylbespuiting beperkte de kolonisatie tot 1200 en 400 propagula per cm² in respectievelijk 1972 en 1973, hetgeen blijkbaar een reductie van de antagonistische capaciteit van de schimmelflora tot gevolg had. In 1974 bereikte de saprofytische kolonisatie op de met water behandelde bladeren, vlak na de bloei, slechts een niveau van 500 propagula per cm², hetgeen niet genoeg bleek om antagonistisch te kunnen zijn.Benomyl had een gedifferentieerd effect op de aantallen fyllosfeerschimmels:Cryptococcus spp. (witte gisten) werden niet nadelig beïnvloed,Sporobolomyces roseus (rode gisten) enCladosporium spp. werden gereduceerd tot minder dan 6% van hun populaties op de contrôle bladeren, enAureobasidium pullulans werd geëlimineerd (Tabel 1). De bacterieflora varieerde enorm per blad en werd niet door benomyl beinvloed.Wanneer later in het seizoen in 1972 en in 1974 de witte gisten op de met benomyl bespoten bladeren populatiedichtheden bereikten van 6500 propagula/cm² of meer, dan was de infectie van deze bladeren vergelijkbaar met die van de met water behandelde bladeren met hogere populatiedichtheden (Fig. 2). Boven een populatiedichtheid van 6500 propagula/cm² bestond er geen correlatie meer tussen de dichtheid van de antagonisten en hun werking.     

Genes Governing Resistance to Puccinia hordei in Thirteen Spring Barley Accessions

ABSTRACT Leaf rust, caused by Puccinia hordei, is an important disease of barley in many parts of the world. In the eastern United States, this disease was effectively controlled for over 20 years through the deployment of cultivars carrying the resistance gene Rph7. Isolates of P. hordei with virulence for Rph7 appeared in this region in the early 1990s rendering barley cultivars with this gene vulnerable to leaf rust infection. From a preliminary evaluation test, 13 accessions from diverse geographic locations possessed resistance to P. hordei isolate VA90-34, which has virulence for genes Rph1, 2, 4, 6, 7, 8, and 11. Each of these 13 accessions was crossed with susceptible cvs. Moore or Larker to characterize gene number and gene action for resistance to P. hordei. Additionally, the 13 accessions were intercrossed and crossed to host differential lines possessing genes Rph3, Rph5, and Rph9 to determine allelic relationships of resistance genes. Seedlings of F(1), F(2), and BC(1)F(1) populations were evaluated in the greenhouse for their reaction to P. hordei isolate VA90-34. Leaf rust resistance in six of the accessions including Collo sib, CR270.3.2, Deir Alla 105, Giza 119, Gloria, and Lenka is governed by a single dominant gene located at or near the Rph3 locus. All accessions for which the gene Rph3 was postulated to govern leaf rust resistance, except for Deir Alla 105, likely possess an allele different than Rph3.c found in Estate based on the differential reaction to isolates of P. hordei. The resistance gene in Grit and Donan is located at or near the Rph9 locus. Alleles at both the Rph3 and Rph9 loci confer resistance in Femina and Dorina. In addition to Rph3, Caroline and CR366.13.2 likely possess a second unknown recessive gene for leaf rust resistance. Resistance in Carre 180 is governed by a recessive gene that is different from all other genes considered in this study. Identification of both known and unique genes conferring leaf rust resistance in the barley germplasm included in this study provides breeding programs with the knowledge and opportunity to assess currently used sources of leaf rust resistance and to incorporate new sources of resistance into their programs.     

Genetics of Stem Rust Resistance in Wheat Cvs. Pasqua and AC Taber

ABSTRACT Canadian wheat cvs. Pasqua and AC Taber were examined genetically to determine the number and identity of stem rust resistance genes in both. The two cultivars were crossed with stem rust susceptible line RL6071, and sets of random F(6) lines were developed from each cross. The F(6) lines, parents, and tester lines with single stem rust resistance genes were grown in a field rust nursery, inoculated with a mixture of stem and leaf rust races, and evaluated for rust resistance. The same wheat lines were tested by inoculation with specific stem rust races in seedling tests to postulate which Sr genes were segregating in the F6 lines. Segregation of F(6) lines indicated that Pasqua had three genes that conditioned field resistance to stem rust and had seedling genes Sr5, Sr6, Sr7a, Sr9b, and Sr12. Leaf rust resistance gene Lr34, which is in Pasqua, was associated with adult-plant stem rust resistance in the segregating F(6) lines. Adult-plant gene Sr2 was postulated to condition field resistance in AC Taber, and seedling genes Sr9b, Sr11, and Sr12 also were postulated to be in AC Taber.     

Identification of Molecular Genetic Markers in Pyrenophora teres f. teres Associated with Low Virulence on 'Harbin' Barley

ABSTRACT Two isolates of the barley net blotch pathogen (Pyrenophora teres f. teres), one possessing high virulence (0-1) and the other possessing low virulence (15A) on the barley cultivar Harbin, were crossed and the progeny of the mating were isolated. Conidia from cultures of the parent and progeny isolates were used as inoculum to determine the inheritance of virulence in the pathogen. Of the 82 progeny tested, 42 exhibited high virulence and 40 exhibited low virulence on 'Harbin' barley. The data support a model in which a single, major gene controls virulence in P. teres f. teres on this barley cultivar (1:1 ratio; chi(2) = 0.05, P = 0.83). Preparations of DNA were made from parental and progeny isolates, and the DNA was subjected to the random amplified polymorphic DNA (RAPD) technique in a search for molecular genetic markers associated with the virulence phenotype. Five RAPD markers were obtained that were associated in coupling with low virulence. The data indicate that the RAPD technique can be used to tag genetic determinants for virulence in P. teres f. teres.     

Specificity of Prehaustorial Resistance to Puccinia hordei and to Two Inappropriate Rust Fungi in Barley

ABSTRACT To elucidate the specificity of prehaustorial resistance to inappropriate rust fungi, we studied two populations of recombinant inbred lines of barley that segregated for partial resistance (PR) to Puccinia hordei and for the resistance to the inappropriate rust species P. recondita f. sp. tritici and P. hordei-murini. PR to P. hordei is prehaustorial and nonhypersensitive, and its level can be assessed accurately by measuring the latent period of the fungus. The resistance to the inappropriate rust species is a combination of prehaustorial (nonhypersensitive) and posthaustorial (hypersensitive) mechanisms. The amount of nonhypersensitive, early abortion of P. recondita f. sp. tritici and P. hordei-murini sporelings reflects the degree of prehaustorial defense to the two inappropriate rust species. All lines showing a long latent period of P. hordei also had a relatively high level of early abortion of the growth of P. recondita f. sp. tritici and P. hordei-murini. This indicates that genes for PR to P. hordei are also effective against these two inappropriate rust species. The reverse was not necessarily true; some lines showing a high level of early abortion of P. recondita f. sp. tritici and P. hordei-murini had a low level of PR to P. hordei. Moreover, lines with a similar level of prehaustorial resistance to P. recondita f. sp. tritici could differ considerably in their prehaustorial resistance to P. hordei-murini. This indicates that genes for prehaustorial resistance may exhibit rust species specificity.     

Expression of Thinopyrum intermedium-Derived Barley yellow dwarf virus Resistance in Elite Bread Wheat Backgrounds

ABSTRACT Resistance to Barley yellow dwarf virus (BYDV) is not found in wheat but is available in a Thinopyrum intermedium translocation (Ti) carried on chromosome 7DL of bread wheat recombinant lines. We used one of those lines (TC14/2*Spear) to introgress the Ti into bread wheat cultivars and to determine the influence of wheat backgrounds, with and without known tolerance to BYDV, on the expression of resistance. Two single and three backcross populations, segregating for the presence of the alien fragment, were tested under field conditions and artificial inoculation with BYDV isolates MAV-Mex and PAV-Mex. Lines containing the fragment were identified using the microsatellite marker gwm37. Tillering, biomass, grain yield, thousand-kernel weight, and seed quality were evaluated in inoculated and noninoculated plots. Resistance was assessed by enzyme-linked immunosorbent assay. In early generations, the alien fragment followed expected Mendelian segregation, whereas in the advanced ones a slight bias against its transmission was observed. No positive nor negative effects of Ti on agronomic performance and quality were found. A significant optical density reduction in individuals carrying the fragment was observed after PAV infection in crosses with lines Anza and Baviacora but not with Milan. In addition, the fragment was associated with a lower frequency of infected plants for both PAV and MAV isolates. The reduced yield loss associated with the presence of the translocation was due largely to the lower infection rate.     

Identification and Mapping of Quantitative Trait Loci Conditioning Resistance to Southern Leaf Blight of Maize Caused by Cochliobolus heterostrophus Race O

ABSTRACT A random set of recombinant inbred (RI) lines (F2:7) derived from the cross of the inbred lines Mo17 (resistant) and B73 (susceptible) were evaluated for resistance to southern leaf blight (SLB) caused by Cochliobolus heterostrophus race O. The RI lines were genotyped at a total of 234 simple sequence repeat, restriction fragment length polymorphism, or isozyme loci. Field plots of the RI lines were inoculated artificially with an aggressive isolate of C. heterostrophus race O in each of two growing seasons in North Carolina. Lines were rated for percent SLB severity two (1996) or three (1995) times during the grain-filling period. Data also were converted to area under the disease progress curve (AUDPC) and analyzed using the composite interval mapping option of the PLABQTL program. When means of disease ratings over years were fitted to models, a total of 11 quantitative trait loci (QTLs) were found to condition resistance to SLB, depending upon which disease ratings were used in the analyses. When the AUDPC data were combined and analyzed over environments, seven QTLs, on chromosomes 1, 2, 3, 4, 7, and 10 were found to come from the resistant parent Mo17. An additional QTL for resistance on chromosome 1 came from the susceptible parent B73. The eight identified QTLs accounted for 46% of the phenotypic variation for resistance. QTL x environment interactions often were highly significant but, with one exception, were the result of differences in the magnitude of QTL effects between years and not due to changes in direction of effects. QTLs on the long arm of chromosome 1 and chromosomes 2 and 3 had the largest effects, were the most consistently detected, and accounted for most of the phenotypic variance. No significant additive x additive epistatic effects were detected. These data support earlier reports of the polygenic inheritance of resistance to SLB of maize.     

Evidence of Cytoplasmic Inheritance of Virulence in Cronartium ribicola to Major Gene Resistance in Sugar Pine

ABSTRACT Tests for Mendelian segregation of virulence and avirulence in Cronartium ribicola, causal agent of white pine blister rust, to a major gene (R) for resistance in sugar pine were made using haploid basidiospore progenies from single diploid telia as inoculum on resistant genotypes. The telia were sampled from a small deme in the Siskyou Mountains of northern California, where a few mature sugar pines known to be Rr genotypes had become infected after withstanding the chronic blister rust epidemic for several decades and where intermediate frequencies of virulence in the ambient basidiospore population were subsequently measured. Infection type on inoculated seedlings with R was qualitative: all progenies of 81 single telia tested over 3 different years were either virulent (compatible) or avirulent (inducing hypersensitive necrosis), never a mixture of both reactions. The complete absence of heterozygotes in the telia population is strong evidence that virulence is not controlled by a nuclear gene. The data are consistent with earlier tests showing that basidiospore inoculum derived from aeciospores isolated from infected Rr trees produced mostly (>90%) virulent reactions on R- seedlings. The evidence indicates that transmission of virulence is uniparental via the cytoplasm of aeciospores. Exchange of spermatia between haploid thalli does not appear to be involved.     

Recessive Genes for Resistance to Puccinia striiformis f. sp. hordei in Barley

ABSTRACT Barley genotypes Abyssinian 14, BBA 2890, Grannelose Zweizeilige, PI 548708, PI 548734, PI 548747, and Stauffers Obersulzer are resistant to all races of Puccinia striiformis f. sp. hordei identified thus far in North America. Astrix, BBA 809, Bigo, Cambrinus, Emir, Heils Franken, Hiproly, I5, Mazurka, Trumpf, and Varunda have specific resistance to certain races, whereas Topper and Steptoe are susceptible to all races. Seedlings of parents and F(1), F(2), and F(3) progeny from crosses of resistant genotypes with Topper and Steptoe were tested for resistance to North American P. striiformis f. sp. hordei races PSH-1, PSH-4, PSH-10, PSH-13, and PSH-20. When tested with PSH-1, one recessive gene was detected in BBA 809, BBA 2890, Bigo, Hiproly, and Grannelose Zweizeilige; two recessive genes were detected in Emir, I5, PI 548708, PI 548734, PI 548747, Varunda, and Astrix; and one dominant gene and one recessive gene were detected in Abyssinian 14 and Stauffers Obersulzer. In tests with PSH-4, one recessive gene was detected in BBA 809, two recessive genes were detected in Trumpf, and two partially recessive genes were detected in Astrix. In tests with PSH-13, one recessive gene was detected in BBA 2890, Grannelose Zweizeilige, I5, and PI 548708 and two recessive genes were detected in Abyssinian 14, Hiproly, PI 548734, and PI 548747. In tests with PSH-20, one recessive gene was detected in Bigo and a recessive gene and a dominant gene were detected in Heils Franken. A gene in Cambrinus for resistance to PSH-10 and a gene in Mazurka for resistance to PSH-20 were dominant on the basis of the response of the first seedling leaf but recessive on the basis of the response of the second leaf. Four different types of epistasis were detected. Information on the number of genes, mode of inheritance, and nonallelic gene interactions should be useful in understanding the host-pathogen interaction and in breeding barley for resistance to stripe rust.     

Diversity and Complexity of Erysiphe graminis f.sp. hordei Collected from Barley Cultivar Mixtures or Barley Plots Treated with a Resistance Elicitor

Powdery mildew populations were analysed to determine the effects of a resistance elicitor and cultivar mixtures on genetic complexity and diversity. Isolations were made from a range of spring barley monocultures and mixtures in a field trial, and characterised for virulence and RAPD profile. In a second trial, isolates were taken from a single mixture from untreated and resistance elicitor-treated areas and from the components of the mixture in monoculture. The mildew population was not only highly heterogeneous for virulence characteristics, but also proved heterogeneous within pathotypes for molecular markers, indicating the major impact of sexual recombination on population structure and the lack of clonal dominance. Various diversity measurements were compared and the value of dissimilarity measurement for revealing genetic distance within a population was highlighted. There was a trend towards increasing complexity as the season progressed, but there was no consistent relationship between cultivar or mixture, disease control treatment, fertiliser treatment, replicate or position in trial, and pathogen genotype. Whilst the resistance elicitor did reduce mildew by 78% in the first trial, and there was no interaction with fertiliser level in its expression, control was substantially less in the second trial. There were no differences between mildew isolates from elicitor and control treatments. It was felt that more effective and consistent resistance elicitors need to be developed before it can be stated that they are unlikely to be eroded by selecting resistant or adapted mildew genotypes.     

Hydrophobin gene expression in the maize pathogen Cochliobolus heterostrophus

Filamentous fungi produce hydrophobins, small proteins localized on the outer surface of their cell walls and involved in growth and development. Hydrophobin gene expression depends on nutrient availability, light, and the activity of conserved signal-transduction pathways. We found four hydrophobins, one class I and three class II family members, in the Cochliobolus heterostrophus genome with high homology to other ascomycete hydrophobins, which present a typical conserved array of cysteines. The expression profile of a selected gene from each class was determined in a series of signaling-deficient mutants. Loss of either of two mitogen-activated protein kinase (MAPK) genes, CHK1 and MPS1, led to decreased hydrophobin class I (CHYD1) gene expression. Mutants in both MAPK genes had easily wettable colonies, but decreased CHYD1 expression was not the sole explanation for this phenotype. A significant elevation of hydrophobin class II (CHYD3) gene expression was measured in the chk1 mutant, suggesting a complex role for MAPK in controlling the expression of these hydrophobins. Similar but less marked tendencies were observed in G-protein α and β subunit loss-of-function mutants; however these showed no alteration in colony hydrophobicity. Overexpression of CHYD1 in the wild-type background caused a change in colony morphology and a small but significant increase in aerial growth. Thus G-protein and MAPK signal transduction influence hydrophobin gene expression and colony hydrophobicity. The connection between colony hydrophobicity and expression of the hydrophobin genes CHYD1 and CHYD3, however, is not one-to-one, indicating that additional factors determine colony-surface properties. The approach of using hydrophobin-overexpression mutants to investigate their role may be generalized to other hydrophobins and small secreted proteins.